title: Efficient viral delivery system for unnatural amino acid mutagenesis in mammalian cells.
abstract: Here we report the development of a baculovirus-based delivery system that enables the efficient incorporation of unnatural amino acids into proteins in mammalian cells. We have exploited the large cargo-capacity (>30 kb) and stability of the double-stranded DNA genome of baculovirus to deliver to a variety of cell types all of the components required to genetically incorporate novel amino acids. These include the engineered tRNA/aminoacyl-tRNA synthetase pair and the nonsense mutant of the target gene. Mammalian cell transduction efficiency of baculovirus was significantly improved by incorporating genetic elements from mammalian viruses. Two polyspecific tRNA/aminoacyl-tRNA synthetase pairs were inserted into this expression system, enabling the site-specific incorporation of a variety of unnatural amino acids with novel chemical and biological properties into proteins.
authors.affiliation: Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA.
authors.forename: Abhishek
authors.initials: A
authors.lastname: Chatterjee
authors.forename: Han
authors.initials: H
authors.lastname: Xiao
authors.forename: Michael
authors.initials: M
authors.lastname: Bollong
authors.forename: Hui-Wang
authors.initials: HW
authors.lastname: Ai
authors.forename: Peter G
authors.initials: PG
authors.lastname: Schultz
mesh_terms: D000596:Amino Acids / Q000378:metabolism*; D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism*; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D004279:DNA, Viral / Q000378:metabolism*; D016503:Drug Delivery Systems / Q000379:methods*; D004926:Escherichia coli; D005347:Fibroblasts; D005818:Genetic Engineering / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics; D051379:Mice; D016296:Mutagenesis / Q000235:genetics*; D012343:RNA, Transfer / Q000378:metabolism*; D061987:Sf9 Cells; D018411:Spodoptera
chemical_list: D000596:Amino Acids; D004279:DNA, Viral; D012343:RNA, Transfer; D000604:Amino Acyl-tRNA Synthetases
keywords: genetic code; nonstandard amino acid; orthogonal; viral vector